Clostridium difficile associated disease (CDAD) is a dangerous emerging infection caused by an anaerobic, spore-forming bacillus that often threatens the health of elderly patients in various healthcare setting following antibiotic treatments of unrelated infections. CDAD symptoms range in severity from asymptomatic carriage, through mild diarrhea, to a more severe pseudomembranous colitis which can be fatal. The epidemiology of CDAD has been changing dramatically. Formerly found almost exclusively as a complication of antimicrobial therapy among the elderly and infirm in inpatient settings, CDAD has been reported increasingly in outpatients, among otherwise healthy individuals including children, and even in the absence of antimicrobial therapy. Deaths attributable to CDAD have quadrupled in the United States from 5.7 per million persons in 1999 to 23.7 per million in 2004. Redelings M D, Sorvillo F, Mascola L. Increase in Clostridium difficile-related mortality rates, United States, 1999-2004. Emerg Infect Dis. 2007; 13:1417-9. Estimates of the cost for treatment for CDAD in the United States have been dramatically revised upward from $1 billion in 2002 to $3.2 billion in 2007 due to a dramatic increase in the number of cases and increasing severity of the disease.
The mechanisms by which C. difficile colonizes the human colon are not established. A surface protein layer has been described which appears to play a role in binding to tissue culture cells in vitro, but its role in vivo has not been established. Calabi, E., Calabi, F., Phillips, A. D. & Fairweather, N. Binding of Clostridium difficile surface layer proteins to gastrointestinal tissues. Infect Immun (2002):70:5770-5778. Enterotoxins A (TcdA) and B (TcdB) are the primary virulence factors of C. difficile. They are exoenzymes that monoglucosylate small Rho-like GTPases, ultimately leading to the disruption of the actin cytoskeleton of colonic intestinal epithelial cells, destruction of tight junction, and apoptosis. Release of cytokines from intoxicated target cells also leads to massive infiltration of neutrophils into damaged tissue regions, a hallmark of pseudomembranous colitis.
Although primary CDAD can usually be successfully treated with metronidazole or vancomycin, metronidazole resistance and refractory infections are becoming increasingly common. Furthermore, many patients suffer recurrent episodes of CDAD, which can be extremely distressing and difficult to manage. An effective vaccine against CDAD is urgently needed for primary and secondary (relapse) prevention. No licensed vaccine is currently available for CDAD. The approach to vaccination that has advanced furthest into clinical trials has targeted only the C. difficile enterotoxins. Toxoid preparations of toxin A and B have completed phase I trials, with both serum free and fecal antibody against both toxins being demonstrated. Kotloff K L, Wasserman S S, Losonsky G A, Thomas W, Jr., Nichols R, Edelman R, Bridwell M, Monath T P. Safety and immunogenicity of increasing doses of a Clostridium difficile toxoid vaccine administered to healthy adults. Infect. Immun. 2001 February; 69(2):988-95. The mechanisms by which serum antibody responses are effective against infection and disease caused by C. difficile are unclear, although it has been proposed that entry of IgG antitoxin from the blood into mucosal tissues of the large bowel or intestinal lumen may prevent toxin binding.
Type W pili (Tfp) or fimbriae are hair-like surface appendages produced by many species of Gram negative bacteria including Pseudomonas aeruginosa, Vibrio cholerae, Neisseria gonorrhoeae, N. meningitidis, Salmonella enterica serovar Typhi (herein designated S. typhi), Legionella pneumophila, enteropathogenic and enterotoxigenic Escherichia coli. Tfp play numerous roles in diverse processes such as cellular adhesion, colonization, twitching motility, biofilm formation, and virulence. Tfp are composed exclusively of primarily of many copies of pilin protein, tightly packed in a helix so that the highly hydrophobic amino-terminus of the pilin is buried in the core of the pilus. Tfp pilins have been used successfully as subunit vaccines for the prevention of several diseases in animals. Lepper A W, Moore L J, Atwell J L, Tennent J M. The protective efficacy of pili from different strains of Moraxella bovis within the same serogroup against infectious bovine keratoconjunctivitis. Vet. Microbiol. 1992; 32:177-87, Lepper A W D, Atwell J L, Lehrbach P R, Schwartzkoff C L, Egerton J R, Tennent J M. The protective efficacy of cloned Moraxella bovis pili in monovalent and multivalent vaccine formulations against experimentally induced infectious bovine keratoconjunctivitis (IBK). Vet. Microbiol. 1995; 45:129-38. Stewart D J, Clark B L, Peterson J E, Emery D L, Smith E F, Griffiths D A, O'Donnell I J. The protection given by pilus and whole cell vaccines of Bacteroides nodosus strain 198 against ovine foot-rot induced by strains of different serogroups. Aust. Vet. J. 1985; 62:153-9. Egerton J R, Cox P T, Anderson B J, Kristo C, Norman M, Mattick J S. Protection of sheep against footrot with a recombinant DNA-based fimbrial vaccine. Vet. Microbiol. 1987; 14:393-409. Recently investigators have discovered that Clostridium perfringens has the genes for and can produce Tfp; similar genes are present in the C. difficile genome. Varga J J, Nguyen V, O'Brien D K, Rodgers K, Walker R A, Melville S B. Type IV pili-dependent gliding motility in the Gram-positive pathogen Clostridium perfringens and other Clostridia. Mol. Microbiol. 2006 November; 62(3):680-94.
There is a need for a multivalent C. difficile subunit vaccine and a diagnostic marker for identification of patients infected with C. difficile. 